KR100618497B1 - Method for extracting quercetin from a propolis - Google Patents

Method for extracting quercetin from a propolis Download PDF

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KR100618497B1
KR100618497B1 KR1020050016781A KR20050016781A KR100618497B1 KR 100618497 B1 KR100618497 B1 KR 100618497B1 KR 1020050016781 A KR1020050016781 A KR 1020050016781A KR 20050016781 A KR20050016781 A KR 20050016781A KR 100618497 B1 KR100618497 B1 KR 100618497B1
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methanol
ratio
chloroform
propolis
quercetin
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이기준
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거창군 (관리부서 : 거창군 농업기술센터)
이기준
박희찬
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Abstract

본 발명은 프로폴리스에서 퀘르세틴을 추출하는 방법에 관한 것으로서, 보다 상세하게는 프로폴리스 에탄올 추출물을 얻는 단계; 상기 프로폴리스 에탄올 추출물에 증류수를 넣고, 클로로포름 및 부탄올을 첨가하여 부탄올 분획물을 분류하는 단계; 상기 부탄올 분획물을 230~400 mesh 크기의 실리카 겔이 충진된 칼럼에 넣은 후, 클로로포름:메탄올의 비율이 99:1로 혼합된 혼합용매에서 메탄올의 비율을 순차적으로 높여 클로로포름:메탄올의 비율이 1:1이 되도록 하여 1차 칼럼 크로마토그래피를 실시하여, 9개의 하위 분획물을 얻는 단계; 클로로포름:메탄올의 비율이 49:1인 시점에서 얻은 하위 분획물 3을 230~400 mesh 크기의 실리카 겔이 충진된 칼럼에 넣은 후, 클로로포름:메탄올의 비율이 49:1로 혼합된 혼합용매에서 메탄올의 비율을 순차적으로 높여 클로로포름:메탄올의 비율이 1:1이 되도록 하여 2차 칼럼 크로마토그래피를 실시하는 단계; 클로로포름:메탄올의 비율이 9:1인 시점에서 퀘르세틴을 얻는 단계를 포함하는 것을 특징으로 한다.The present invention relates to a method for extracting quercetin from propolis, and more particularly, obtaining a propolis ethanol extract; Distilled water is added to the propolis ethanol extract, and chloroform and butanol are added to classify the butanol fraction; After putting the butanol fraction in a column filled with silica gel having a size of 230 to 400 mesh, the ratio of chloroform: methanol is increased by sequentially increasing the ratio of methanol in a mixed solvent having a chloroform: methanol ratio of 99: 1. Performing primary column chromatography to 1 to obtain 9 subfractions; Sub-fraction 3 obtained at the time when the ratio of chloroform: methanol was 49: 1 was placed in a column filled with silica gel having a size of 230 to 400 mesh, and then methanol was mixed in a mixed solvent having a chloroform: methanol ratio of 49: 1. Sequentially increasing the ratio so that the ratio of chloroform: methanol is 1: 1 to perform secondary column chromatography; Obtaining quercetin at a chloroform: methanol ratio of 9: 1.

퀘르세틴, 칼럼 크로마토그래피, 프로폴리스, 플라보노이드 Quercetin, column chromatography, propolis, flavonoids

Description

프로폴리스에서 퀘르세틴을 추출하는 방법{Method for extracting quercetin from a propolis}Method for extracting quercetin from a propolis

도 1은 본 발명에 의한 퀘르세틴의 IR 스펙트럼 측정 결과를 나타낸 도면,1 is a view showing an IR spectrum measurement result of quercetin according to the present invention,

도 2는 본 발명에 의한 퀘르세틴의 질량 스펙트럼 측정 결과를 나타낸 도면,2 is a view showing a mass spectrum measurement result of quercetin according to the present invention;

도 3은 본 발명에 의한 퀘르세틴의 1H-NMR 스펙트럼 측정 결과를 나타낸 도면,3 is a view showing a 1 H-NMR spectrum measurement results of quercetin according to the present invention,

도 4는 본 발명에 의한 퀘르세틴의 13C-NMR 스펙트럼 측정 결과를 나타낸 도면,Figure 4 shows the 13 C-NMR spectrum measurement results of quercetin according to the present invention,

도 5는 본 발명에 의한 퀘르세틴의 13C-1H COSY(HMQC) 스펙트럼 측정 결과를 나타낸 도면,5 is a view showing a 13 C- 1 H COSY (HMQC) spectrum measurement results of quercetin according to the present invention,

도 6은 본 발명에 의한 퀘르세틴의 1H-1H COSY 스펙트럼 측정 결과를 나타낸 도면,Figure 6 is a view of the 1 H- 1 H COSY spectrum measurement result of quercetin according to the invention,

도 7은 본 발명에 의한 퀘르세틴의 HMBC 스펙트럼 측정 결과를 나타낸 도면이다.7 shows the results of measuring the HMBC spectrum of quercetin according to the present invention.

본 발명은 프로폴리스에서 퀘르세틴(quercetin)을 추출하는 방법에 관한 것으로서, 보다 상세하게는 프로폴리스로부터 칼럼 크로마토그래피 방법을 이용하여 항암능력이 있는 퀘르세틴을 추출하는 방법에 관한 것이다.The present invention relates to a method of extracting quercetin from propolis, and more particularly, to a method of extracting quercetin having anticancer ability from columnar methods from propolis.

프로폴리스는 벌집에서 얻어지는 지용성 복합체로서, 꿀벌들이 벌집내에 들어있는 꿀을 보존하기 위하여 벌집의 틈새를 튼튼하게 만들기 위해 사용되는 물질로, 벌이 꽃, 잎, 봉오리 등으로부터 수집하는 왁스와 수지(resin)물질을 벌 자신의 침샘 분비물과 혼합하여 만드는 수지성, 점착성, 고무상의 물질이다.Propolis is a fat-soluble complex obtained from honeycomb, a substance used by bees to strengthen the honeycomb crevices to preserve the honey contained in the honeycomb. Wax and resin that bees collect from flowers, leaves, buds, etc. It is a resinous, tacky, rubbery substance made by mixing a substance with its own salivary glands.

프로폴리스는 계피나 바닐라 등을 혼합한 것과 같은 독특한 향기가 나며, 50~55 중량%의 수지 및 방향유 물질, 25~35 중량%의 밀납, 10 중량%의 정유, 5 중량%의 화분으로 이루어져 있고, 이외에도 아미노산, 유기산, 화분, 미량원소(알루미늄, 칼슘, 규소, 철, 구리, 망간, 아연, 스트론튬), 비타민, 플라보노이드, 프로비타민, 항균물질, 효소 등 다양한 성분을 5 중량% 함유하고 있다.Propolis has a unique scent, such as a mixture of cinnamon and vanilla, and consists of 50-55% by weight of resin and aromatic substances, 25-35% by weight of beeswax, 10% by weight of essential oils, 5% by weight of pollen. In addition, it contains 5% by weight of various components such as amino acids, organic acids, pollen, trace elements (aluminum, calcium, silicon, iron, copper, manganese, zinc, strontium), vitamins, flavonoids, provitamins, antibacterial substances, enzymes, and the like.

프로폴리스의 생리활성에 관하여는 이미 많은 연구가 진행되어 항암, 항균, 항박테리아, 항바이러스, 항염증, 항진균성, 항산화 및 알레르기성 피부염 치료에도 매우 높은 활성을 나타내고 있다. 이와 같이 프로폴리스가 다양한 생리활성을 가지는 주된 이유는, 프로폴리스의 주성분인 플라보노이드에 의한 것으로 이미 입증되어 있다.Many studies have been conducted on the physiological activity of propolis, which has been shown to be highly effective in the treatment of anticancer, antibacterial, antibacterial, antiviral, anti-inflammatory, antifungal, antioxidant and allergic dermatitis. As such, the main reason why propolis has various physiological activities has already been demonstrated by flavonoids, which are the main components of propolis.

프로폴리스에는 약 20여 종류의 플라보노이드가 함유되어 있으며, 신남산 (cinnamic acid) 및 카페인산 에스테르(caffeic acids ester)를 비롯한 많은 종류의 플라보노이드는 인체 내 백혈구나 림프사이트를 자극하여 인터페론 생성을 활성화하고, 체내 면역력을 증강시키는 동시에 항산화, 항염, 항알레르기 효과가 매우 강력한 물질로 알려져 있다.Propolis contains about 20 kinds of flavonoids, and many types of flavonoids, including cinnamic acid and caffeic acids esters, stimulate the white blood or lymphocytes in the body to activate interferon production. In addition, it is known to enhance the body's immunity and at the same time have strong antioxidant, anti-inflammatory and anti-allergic effects.

특히, 플라보노이드의 한 종류인 퀘르세틴(quercetin)은 식품 중에 존재하는 플라보노이드 중에서 대표적인 것으로 항산화 작용을 하여 노화를 억제하고 동맥경화를 방지할 수 있는 효능을 가지고 있는 물질이다.In particular, one of the flavonoids quercetin (quercetin) is a representative of the flavonoids present in the food is an substance that has the effect of inhibiting aging and anti-arteriosclerosis by the antioxidant action.

본 발명의 목적은 프로폴리스로 부터 칼럼 크로마토그래피 기법을 이용하여 퀘르세틴을 효율적으로 추출하는 방법을 제공하고자 하는 것이다.It is an object of the present invention to provide a method for efficiently extracting quercetin from propolis using column chromatography techniques.

상기 목적을 달성하기 위한 본 발명의 프로폴리스에서 퀘르세틴을 추출하는 방법은 프로폴리스 에탄올 추출물을 얻는 단계; 상기 프로폴리스 에탄올 추출물에 증류수를 넣고, 클로로포름 및 부탄올을 첨가하여 부탄올 분획물을 분류하는 단계; 상기 부탄올 분획물을 230~400 mesh 크기의 실리카 겔이 충진된 칼럼에 넣은 후, 클로로포름:메탄올의 비율이 99:1로 혼합된 혼합용매에서 메탄올의 비율을 순차적으로 높여 클로로포름:메탄올의 비율이 1:1이 되도록 하여 1차 칼럼 크로마토그래피를 실시하여, 9개의 하위 분획물을 얻는 단계; 클로로포름:메탄올의 비율이 49:1인 시점에서 얻은 하위 분획물 3을 230~400 mesh 크기의 실리카 겔이 충진된 칼럼에 넣은 후, 클로로포름:메탄올의 비율이 49:1로 혼합된 혼합용매에서 메탄올의 비 율을 순차적으로 높여 클로로포름:메탄올의 비율이 1:1이 되도록 하여 2차 칼럼 크로마토그래피를 실시하는 단계; 클로로포름:메탄올의 비율이 9:1인 시점에서 퀘르세틴을 얻는 단계를 포함하는 것을 특징으로 한다.Method for extracting quercetin from the propolis of the present invention for achieving the above object comprises the steps of obtaining a propolis ethanol extract; Distilled water is added to the propolis ethanol extract, and chloroform and butanol are added to classify the butanol fraction; After putting the butanol fraction in a column filled with silica gel having a size of 230 to 400 mesh, the ratio of chloroform: methanol is increased by sequentially increasing the ratio of methanol in a mixed solvent having a chloroform: methanol ratio of 99: 1. Performing primary column chromatography to 1 to obtain 9 subfractions; Sub-fraction 3 obtained at the time when the ratio of chloroform: methanol was 49: 1 was placed in a column filled with silica gel having a size of 230 to 400 mesh, and then methanol was mixed in a mixed solvent having a chloroform: methanol ratio of 49: 1. Sequentially increasing the ratio so that the ratio of chloroform: methanol is 1: 1 to perform secondary column chromatography; Obtaining quercetin at a chloroform: methanol ratio of 9: 1.

여기에서, 상기 프로폴리스 에탄올 추출물은 프로폴리스에 70% 에탄올 용매를 첨가하고, 상온에서 3일 간격으로 3시간씩 3회 반복 추출한 후 농축하여 얻어지는 것을 특징으로 한다.Here, the propolis ethanol extract is characterized in that it is obtained by adding 70% ethanol solvent to the propolis, and extracted three times at three times intervals at room temperature three times at room temperature and then concentrated.

이하, 본 발명을 보다 상세히 설명하면 다음과 같다.Hereinafter, the present invention will be described in more detail.

본 발명에서 사용되는 프로폴리스는 경남 거창 일원에서 채취한 국내산 프로폴리스로서, 우선 상기 프로폴리스 2㎏에 70% 에탄올 용매 4ℓ를 첨가하고, 상온에서 3일 간격으로 3시간씩 3회 반복 추출한 후, 회전증발기(rotary evaporator)로 농축하여 약 300g의 프로폴리스 에탄올 추출물을 얻을 수 있었다.The propolis used in the present invention is a domestic propolis collected from Gyeongnam Geochang, first, 4L of 70% ethanol solvent is added to 2 kg of the propolis, and extracted three times at 3 days intervals at room temperature for 3 hours. Concentration by rotary evaporator (rotary evaporator) to obtain about 300g of propolis ethanol extract.

상기와 같은 방법으로 얻은 상기 프로폴리스 에탄올 추출물에 증류수를 넣고 클로로포름(chloroform) 및 부탄올(butanol)을 첨가하고 분류하여, 클로로포름(CHCl3) 분획물, 부탄올(BuOH) 분획물 및 물 분획물을 각각 얻었으며, 이중 가장 생리활성이 뛰어난 부탄올 분획물을 이용하였다.Distilled water was added to the propolis ethanol extract obtained by the above method, chloroform and butanol were added and sorted to obtain a chloroform (CHCl 3 ) fraction, butanol (BuOH) fraction and water fraction, respectively. Among them, the butanol fraction having the most physiological activity was used.

상기 부탄올 분획물을 230~400 mesh 크기의 실리카 겔(silica gel)이 충진된 칼럼(column)에 넣은 후, 클로로포름:메탄올의 비율이 99:1로 혼합된 용매에서 상기 메탄올의 비율을 순차적으로 늘려주어, 클로로포름:메탄올의 비율이 1:1이 되도록 하여 1차 칼럼 크로마토그래피(column chromatography)를 실시한다. The butanol fraction was placed in a column filled with silica gel having a size of 230 to 400 mesh, and the ratio of methanol was sequentially increased in a solvent in which the ratio of chloroform: methanol was mixed at 99: 1. , Primary column chromatography is performed so that the ratio of chloroform: methanol is 1: 1.

이때, 용출액은 250㎖씩 회수하였으며, 상기 부탄올 분획물으로 부터 9개의 하위 분획물(subfraction)을 얻을 수 있었다. 여기에서, 분획물의 분류기준은 프리코티드 티엘씨 플레이트(precoated TLC plate)를 사용하여 상승 1차원법으로 전개시킨 후, UV254㎚ 조사와, 용출액을 10% 황산으로 발색시키면서 나타나는 티엘씨 패턴(TLC pattern)에 따른 것으로서, 상기 9개의 하위 분획물(subfraction)은 분획물 1 부터 분획물 9 까지 지정되었다.At this time, the eluate was recovered by 250 ml, and nine subfractions were obtained from the butanol fraction. Herein, the classification criteria of the fractions were developed by a synergistic one-dimensional method using a precoated TLC plate, followed by UV254 nm irradiation, and the eluate with the 10% sulfuric acid. According to the pattern), the nine subfractions were designated from fraction 1 to fraction 9.

상기 9개의 하위 분획물 중에서 UV254㎚에서 활동하며, 크로마토그래피 기법에서 Rf값이 0.28인 화합물을 분리하기 위하여, 클로로포름:메탄올의 비율이 49:1인 시점에서 얻은 하위 분획물 3을 230~400 mesh 크기의 실리카 겔(silica gel)이 충진된 칼럼(column)에 넣은 후, 클로로포름:메탄올의 비율이 49:1로 혼합된 용매에서 상기 메탄올의 비율을 순차적으로 늘려주어, 클로로포름:메탄올의 비율이 1:1이 되도록 하여 2차 칼럼 크로마토그래피(column chromatography)를 실시한다.In order to separate the compound having an Rf value of 0.28 in the chromatographic technique, the subfraction 3 obtained at the time of chloroform: methanol ratio of 49: 1 among the nine subfractions was operated at UV254 nm. After the silica gel was filled in a column, the ratio of chloroform: methanol was sequentially increased in a solvent in which the ratio of chloroform: methanol was 49: 1, and the ratio of chloroform: methanol was 1: 1. Second column chromatography is performed so that it may become.

최종적으로 클로로포름:메탄올의 비율이 9:1 되는 시점에서 용리되는 노란색 분말의 퀘르세틴을 얻었으며, 상기 퀘르세틴의 순도확인은 프리코티드 티엘씨 (precoated TLC) 1차원 상승법으로 확인하였다.Finally, when the ratio of chloroform: methanol was 9: 1, a yellow powder of quercetin eluted was obtained, and the purity of the quercetin was confirmed by precoated TLC one-dimensional ascension method.

상기와 같은 방법으로 얻은 퀘르세틴은 플라보놀(flavonol)골격의 C3, C5, C7, C3' 및 C4' 위치에 OH기를 가지고 있는 3',4',5,7-tetrahydroxy-flavonol 이며, 분자량은 302이다.Quercetin obtained by the above method is 3 ', 4', 5,7-tetrahydroxy-flavonol having an OH group at the C3, C5, C7, C3 'and C4' positions of the flavonol skeleton, and the molecular weight is 302 to be.

또한, 퀘르세틴의 분자식은 C15H10O7이며 아래 화학식 1과 같다.In addition, the molecular formula of quercetin is C 15 H 10 O 7 It is represented by the following formula (1).

Figure 112005010862590-pat00001
Figure 112005010862590-pat00001

본 발명에 따른 추출방법으로 추출된 상기 퀘르세틴에 관해 보다 상세히 설명하면 다음과 같다.The quercetin extracted by the extraction method according to the present invention will be described in more detail as follows.

실험예Experimental Example 1: IR 스펙트럼(spectrum) 측정 1: IR spectrum measurement

본 실험예에서 측정한 퀘르세틴의 IR 스펙트럼 측정 결과가 도 1에 도시되어 있다.IR spectrum measurement results of quercetin measured in the present experimental example are shown in FIG. 1.

도 1에 나타난 바와 같이, 퀘르세틴은 3401cm-1에서 하이드록실(hydroxyl)기, 1662cm-1에서 카보닐 그룹(carbonyl group), 1611cm-1 및 1517cm-1에서 아로마틱(aromatic) C=C의 흡수 피크(peak)를 나타냄을 확인할 수 있었다.As shown in Fig. 1, quercetin is hydroxyl (hydroxyl) group, 1662cm -1 in absorption of carbonyl group (carbonyl group), 1611cm -1 and aromatic (aromatic) C = C peak at 1517cm -1 3401cm -1 in (peak) was confirmed.

실험예Experimental Example 2: 질량 스펙트럼(mass spectrum) 측정 2: Mass spectrum measurement

본 실험예에서 측정한 퀘르세틴의 질량 스펙트럼 측정 결과가 도 2에 도시되어 있다.The mass spectrum measurement result of quercetin measured in this experimental example is shown in FIG. 2.

도 2에 나타난 바와 같이, 퀘르세틴은 m/z=302인 지점에서 분자 이온 피크(molecular ion peak)를 나타냄을 확인할 수 있었다.As shown in FIG. 2, it was confirmed that quercetin exhibits a molecular ion peak at the point of m / z = 302.

실험예Experimental Example 3:  3: 1One H-NMR 스펙트럼(spectrum) 측정H-NMR spectrum measurement

본 실험예에서 측정한 퀘르세틴의 1H-NMR 스펙트럼 측정 결과가 도 3에 도시되어 있다. 1 H-NMR spectrum measurement results of quercetin measured in the present experimental example are shown in FIG. 3.

도 3에 나타난 1H-NMR 스펙트럼의 적분비로부터 퀘르세틴의 수소수가 10개임을 확인할 수 있었다.From the integral ratio of the 1 H-NMR spectrum shown in Figure 3 it can be confirmed that the number of hydrogen of quercetin 10.

또한, 케미컬 쉬프트(chemical shift)값 및 결합 상수(coupling constant)값으로부터 A 고리(A-ring)의 메틴(methine)기의 메타 결합(meta coupling)이 δ6.22(1H, d, J=2.0Hz, H-6)과 δ6.44(1H, d, J=2.0Hz, H-8)에서 나타났고, B 고리(B-ring)의 H-2', H-6' 및 H-5'의 시그널(signal)이 각각 δ7.71(1H, d, J=2.2Hz, H-2'), δ7.57(1H, dd, J=8.5, 2.2Hz, H-6') 및 δ6.92(1H, d, J=8.5Hz, H-5')에서 메타(meta)와 오르토 결합(ortho coupling)을 나타냄을 확인할 수 있었다.In addition, the meta coupling of the methine group of the A-ring from the chemical shift value and the coupling constant value is δ6.22 (1H, d, J = 2.0). Hz, H-6) and δ6.44 (1H, d, J = 2.0 Hz, H-8), and H-2 ', H-6' and H-5 'of the B-ring. Signals of δ7.71 (1H, d, J = 2.2Hz, H-2 '), δ7.57 (1H, dd, J = 8.5, 2.2Hz, H-6') and δ6.92, respectively. At (1H, d, J = 8.5 Hz, H-5 ′), meta and ortho coupling were shown.

실험예Experimental Example 4:  4: 1313 C-NMR 스펙트럼(spectrum) 측정C-NMR Spectrum Measurement

본 실험예에서 측정한 퀘르세틴의 13C-NMR 스펙트럼 측정 결과가 도 4에 도시되어 있다. 13 C-NMR spectrum measurement results of quercetin measured in the present experimental example are shown in FIG. 4.

도 4에 나타난 바와 같이, 퀘르세틴의 13C-NMR 스펙트럼에서는 총 15개의 탄소 시그널(carbon signal)이 확인되었으며, δ174.7의 피크 시그널(peak signal)은 카르보닐(carbonyl)기 임을 확인할 수 있었다.As shown in FIG. 4, a total of 15 carbon signals were identified in the 13 C-NMR spectrum of quercetin, and a peak signal of δ 174.7 was confirmed to be a carbonyl group.

실험예Experimental Example 5:  5: 1313 C-C- 1One H COSY(H COSY ( HMQCHMQC ) 스펙트럼(spectrum) 측정Spectrum measurement

본 실험예에서 측정한 퀘르세틴의 13C-1H COSY(HMQC) 스펙트럼 측정 결과가 도 5에 도시되어 있으며, 이를 통해 퀘르세틴을 구성하는 각 탄소에 결합된 수소와의 상관관계(cross peak)를 확인할 수 있었다. 13 C- 1 H COSY (HMQC) spectrum measurement results of quercetin measured in the present experimental example is shown in Figure 5, through this to confirm the cross (peak) cross-linked hydrogen with each carbon constituting the quercetin Could.

실험예Experimental Example 6:  6: 1One H-H- 1One H COSY 스펙트럼(spectrum) 측정H COSY spectrum measurement

본 실험예는 인접한 수소사이의 상관관계를 확인하기 위하여 실시하였으며, 본 실험예에서 측정한 퀘르세틴의 1H-1H COSY 스펙트럼 측정 결과가 도 6에 도시되어 있다.This experimental example was carried out to determine the correlation between the adjacent hydrogen, the 1 H- 1 H COSY spectrum measurement result of a measurement of quercetin in this experimental example is shown in FIG.

이를 통하여, H-2'(δ7.71), H-5'(δ6.92), H-6'(δ7.57)은 서로 상관관계가 있음을 확인할 수 있었고, H-6(δ6.22)와 H-8(δ7.57)의 상관관계 또한 확인할 수 있었다.Through this, it was confirmed that H-2 '(δ7.71), H-5' (δ6.92), and H-6 '(δ7.57) correlated with each other, and H-6 (δ6.22 ) And H-8 (δ7.57) were also identified.

즉, H-2'(δ7.71)와 H-6'(δ7.57)의 수소는 서로 메타 결합(meta coupling)을 하고, H-6'(δ7.57)와 H-5'(δ6.92)의 수소는 서로 오르토 결합(ortho coupling)을 하고, H-8(δ7.57)와 H-6(δ6.22)의 수소는 서로 메타 결합(meta coupling)을 함을 확인할 수 있었다.That is, hydrogen of H-2 '(δ7.71) and H-6' (δ7.57) is meta-coupled with each other, and H-6 '(δ7.57) and H-5' (δ6) Hydrogen of .92) is ortho-coupled with each other, and hydrogen of H-8 (δ7.57) and H-6 (δ6.22) is meta-coupled with each other.

실시예Example 7:  7: HMBCHMBC 스펙트럼(spectrum) 측정 Spectrum Measurement

본 실험예는 퀘르세틴의 보다 구체적인 구조동정 및 제 4차 탄소에 대한 정보를 얻기 위해 실시하였으며, 본 실험예에서 측정한 퀘르세틴의 HMBC 스펙트럼 측정 결과가 도 7에 도시되어 있다.This experimental example was carried out to obtain more detailed structure identification of quercetin and information on the fourth carbon, and the HMBC spectrum measurement result of quercetin measured in this experimental example is shown in FIG. 7.

이를 통하여, δ120.9, δ145.7 및 δ143.9의 4차 탄소 시그널(signal)들은 각각 플라보놀(flavonol)의 B 고리(B-ring)에 위치한 C-1', 3' 및 4'의 시그널임을 알 수 있었고, δ134.6의 탄소 시그널은 C-3임을 알 수 있었다.Through this, the quaternary carbon signals of δ 120.9, δ 145.7 and δ 143.9 are respectively represented by C-1 ′, 3 ′ and 4 ′ located in the B-ring of flavonol. It was found that the signal, the carbon signal of δ 134.6 was found to be C-3.

본 발명에 따른 프로폴리스에서 퀘르세틴을 추출하는 방법에 의하면, 기존의 양파 등의 식품에서 얻을 수 있었던 항암작용 및 동맥경화 방지의 효능을 가진 퀘르세틴을 프로폴리스로부터 칼럼 크로마토그래피 기법을 이용하여 효율적으로 획득할 수 있다는 장점이 있다.According to the method of extracting quercetin from the propolis according to the present invention, quercetin having an anti-cancer effect and atherosclerosis prevention effect obtained from foods such as onions can be efficiently obtained from the propolis using column chromatography. The advantage is that you can.

Claims (2)

프로폴리스 에탄올 추출물을 얻는 단계;Obtaining propolis ethanol extract; 상기 프로폴리스 에탄올 추출물에 증류수를 넣고, 클로로포름 및 부탄올을 첨가하여 부탄올 분획물을 분류하는 단계;Distilled water is added to the propolis ethanol extract, and chloroform and butanol are added to classify the butanol fraction; 상기 부탄올 분획물을 230~400 mesh 크기의 실리카 겔이 충진된 칼럼에 넣은 후, 클로로포름:메탄올의 비율이 99:1로 혼합된 혼합용매에서 메탄올의 비율을 순차적으로 높여 클로로포름:메탄올의 비율이 1:1이 되도록 하여 1차 칼럼 크로마토그래피를 실시하여, 9개의 하위 분획물을 얻는 단계;After putting the butanol fraction in a column filled with silica gel having a size of 230 to 400 mesh, the ratio of chloroform: methanol is increased by sequentially increasing the ratio of methanol in a mixed solvent having a chloroform: methanol ratio of 99: 1. Performing primary column chromatography to 1 to obtain 9 subfractions; 클로로포름:메탄올의 비율이 49:1인 시점에서 얻은 하위 분획물 3을 230~400 mesh 크기의 실리카 겔이 충진된 칼럼에 넣은 후, 클로로포름:메탄올의 비율이 49:1로 혼합된 혼합용매에서 메탄올의 비율을 순차적으로 높여 클로로포름:메탄올의 비율이 1:1이 되도록 하여 2차 칼럼 크로마토그래피를 실시하는 단계;Sub-fraction 3 obtained at the time when the ratio of chloroform: methanol was 49: 1 was placed in a column filled with silica gel having a size of 230 to 400 mesh, and then methanol was mixed in a mixed solvent having a chloroform: methanol ratio of 49: 1. Sequentially increasing the ratio so that the ratio of chloroform: methanol is 1: 1 to perform secondary column chromatography; 클로로포름:메탄올의 비율이 9:1인 시점에서 퀘르세틴을 얻는 단계;Obtaining quercetin at a chloroform: methanol ratio of 9: 1; 를 포함하는 것을 특징으로 하는 프로폴리스에서 퀘르세틴을 추출하는 방법.Method of extracting quercetin from propolis comprising a. 제 1항에 있어서,The method of claim 1, 상기 프로폴리스 에탄올 추출물은, The propolis ethanol extract, 프로폴리스에 70% 에탄올 용매를 첨가하고, 상온에서 3일 간격으로 3시간씩 3회 반복 추출한 후 농축하여 얻어지는 것을 특징으로 하는 프로폴리스에서 퀘르세 틴을 추출하는 방법.A method of extracting quercetin from propolis, which is obtained by adding 70% ethanol solvent to propolis, and extracting three times at three days intervals at room temperature for three hours.
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20170116688A (en) * 2016-04-12 2017-10-20 대한민국(농촌진흥청장) Method of simultaneous analysing for para-coumaric acid and trans-cinnamic acid in propolis using ultra-high performance liquid chromatography

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH05271031A (en) * 1991-07-03 1993-10-19 Nippon Puroporisu Kk Production of extract of propolis
JPH07330596A (en) * 1994-06-01 1995-12-19 Eiken Chem Co Ltd Antitumor agent
JP2000103789A (en) 1998-09-28 2000-04-11 Mikako Hirota Physiological activity of benzopyran derivative derived from propolis

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH05271031A (en) * 1991-07-03 1993-10-19 Nippon Puroporisu Kk Production of extract of propolis
JPH07330596A (en) * 1994-06-01 1995-12-19 Eiken Chem Co Ltd Antitumor agent
JP2000103789A (en) 1998-09-28 2000-04-11 Mikako Hirota Physiological activity of benzopyran derivative derived from propolis

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20170116688A (en) * 2016-04-12 2017-10-20 대한민국(농촌진흥청장) Method of simultaneous analysing for para-coumaric acid and trans-cinnamic acid in propolis using ultra-high performance liquid chromatography
KR101883833B1 (en) 2016-04-12 2018-08-01 대한민국 Method of simultaneous analysing for para-coumaric acid and trans-cinnamic acid in propolis using ultra-high performance liquid chromatography

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